Cells of the innate immune system express pattern recognition receptors (PRR), or Toll-like receptors (TLR), sensing pathogen-associated molecular patterns (PAMP). TLR engagement leads to the activation of intracellular signaling pathways and NFkB transcription factor involved in the activation of a variety of pro-inflammatory genes presumably via a universal adaptor, MyD88. We hypothesize that Vav proteins are crucial mediators of macrophage and B cell responses to bacterial endotoxins by regulating the activity of Pyk2 kinase and actin cytoskeletal changes. Our preliminary observations show that murine B cells and macrophages deficient in all 3 Vav-family proteins fail to respond to bacterial endotoxin stimulation in vitro. To determine the requirement for Vav proteins in signaling downstream of TLRs in mediating inflammatory responses will use Vav1/2/3-deficient mice and to develop in vivo assays for structure/function analyses of Vav. Specifically we propose: 1. SPECIFIC AIM 1. To determine the requirement for Vav proteins in responses to endotoxins. SPECIFIC AIM 2. To determine the structural basis of Vav function in Toll-like receptor signaling.